Synthesis and evaluation of an ¹⁸F-labeled sulfonamide inhibitor for imaging carbonic anhydrase IX expression in cancers

Objectives Carbonic anhydrase IX (CA-IX), a transmembrane protein, is overexpressed in a variety of cancers under hypoxic stress. U-104, a sulfonamide derivative and potent CA-IX inhibitor, has been shown to effectively inhibit primary breast cancer growth and distant metastasis in xenograft models. Herein, we synthesized and evaluated [¹⁸F]U-104 as a potential PET tracer for imaging CA-IX expression in cancers.

Methods Ki of U-104 to CA isoenzymes was measured via CA catalyzed CO₂ hydration assays. [¹⁸F]U-104 was synthesized in three steps: nucleophilic substitution of 1,4-dinitrobenzene with [¹⁸F]]fluoride to form 1-[¹⁸F]fluoro-4-nitrobenzene, reduction to yield 4-[¹⁸F]fluoroaniline, and final coupling with 4-nitrophenyl 4-sulfamoylcarbanilate to generate [¹⁸F]U-104. Stability of [¹⁸F]U-104 was determined in mouse plasma. Biodistribution study was performed using NODSCID/IL2RKO mice bearing CA-IX expressing HT-29 colorectal tumors. The expression of CA-IX in HT-29 tumors was confirmed by immunostaining.

Results The Ki of U-104 for CA isoenzymes I, II, and IX were 5,080, 96 and 45 nM, respectively. [¹⁸F]U-104 was obtained in 6% overall decay-corrected yield in 150 min with > 98% radiochemical purity and 15.1-19.8 Ci/μmol specific activity at EOS. [¹⁸F]U-104 was stable in mouse plasma with negligible decomposition after 2 h incubation at 37 °C. Biodistribution study showed that the radioactivity was excreted via both renal and hepatobiliary pathways. Tumor uptake (%ID/g) of [¹⁸F]U-104 at 1 h p.i. was 0.83 ± 0.06 which was lower than blood (13.9 ± 0.43), muscle (1.19 ± 0.20), and major organs except brain (0.16 ± 0.01).

Conclusions High blood uptake in mice is likely due to the binding of [¹⁸F]U-104 to CA-II in erythrocytes. Because of minimal tumor uptake relative to normal tissues, [¹⁸F]U-104 is not suitable for use for CA-IX targeted imaging.

Research Support This work was supported by the Canadian Institutes of Health Research (CIHR).